A novel ELISA for the evaluation of the classical pathway of complement

Abstract

Assessment of the overall function of the classical pathway of complement is traditionally performed by the hemolytic titration assay CH50. In the present study, we established a novel method for the quantitation of complement activity by measuring the deposition of C1q, C4, C3 and C9 on solid-phase IgM by an enzyme-linked immunosorbent assay (ELISA). Using the CH50 method as the reference, C9 deposition values displayed a sensitivity of 96.3% and a specificity of 99.4% in sera from patients with a variety of diseases. For C3, the sensitivity was 91.3% and the specificity 100%, for C4, the values were 95% and 100%, and for C1q, the corresponding values were 52.9% and 98.9%. A close correlation was found between CH50 values below 30 U/ml and the deposition of C9 ( r=0.92), C3 ( r=0.91) and C4 ( r=0.92). In two patients with postinfectious glomerulonephritis normal C4 and C1q deposition was accompanied by decreased C3 and C9 deposition reflecting complement activation predominantly through the alternative pathway. In contrast, in two patients with complete C2 deficiency the deposition of C3 and C9 was undetectable together with normal C4 deposition values. Furthermore, in two patients with hereditary C1-inhibitor deficiency distinctly increased C1q deposition was accompanied by decreased C4 deposition values. In conclusion, the determination of complement deposition by ELISA represents a novel, quantitative method for the evaluation of complement activity. The measurement of C9 deposition alone or in combination with further complement proteins makes this ELISA a valuable tool for assessing the degree and level of complement consumption as well as localizing the missing protein in the case of complement deficiencies.